(1) Axnic cultivation of Entamoeba dispar : Hydrogen peroxide treated Crithidia fasciculata cartainly promoted the growth of E.dispar in vitro ; however, recent NMR analysis indicated that the flagellate still had signals due to amino acids and some organic acids suggesting that such flagellates still maintained metabolic activities. Subsequent trial using the flagellates treated at 56ﾟC,30 min followed by H_2O_2 treatment resulted in complete loss of the signals on NRM.Such sterilized and metabolically inactive flagellates had a favorale promoting activity for the in vitro growth of E.dispar, which is consistent with the definition of axnic cultivation. Moreover, we have recently developed the complete axnic medium without any culture supplement, which enabled us to mainatain 3 strains under such conditions.(2) Ultrastructural difference : In E.dispar, numerous glycogen particles were detected primarily on the periphery of cytoplasm surrounding undeveloped cellular organelles at the c
… Moreenter, while the particles were dispersed evenly throughout the cytoplasm in E.histolytica grown under the same conditions.(3) Biochemical studies : The mst striking difference was the amount of glycogen. E.dispar trophozoite contained 8^-9 times as much glycogen per mg protein as E.histolytica. In addition, fructose well promoted the growth of E.dispar, while glucose was better to support the growth of E.histolytica, which enabled us to characterize hexokinase and glucokinase in E.dispar and E.histolytica respectiely.(4) Genetic studies : Now we arefocusing our efforts on sulfur metabolism of E.histolytica. We were able to clone cDNA for L-cystein synthase, which has been detected only in plants ans bacteria. This gene was located next to NADP^+-dependent alcohol dehyrogenase gene. Both of these proteins have been considered as major antigens related with chronic colitis. The reason for this is under current investigation. Moreover, we did not detect m-RNA for L-cysten synthase, the reason of which is also being studied.(5) Pathogenicity and virulence of E.dispar : Axnic strains of E.dispar were inculated into liver of newborn hamsters to assess the virulence ; however, no nistopathological changes could be found. Immunocompromized animals were induced by injecting steroid repeatedly into hamsters, which did not lead to formation of abscess lesion in the liver by E.dispar, sugegsting virulence of E.dispar should be negligible. Less